Staphylococcus aureus is a common human commensal bacterium that is capable of causing a wide range of serious infections, accounting for billions of dollars in health care costs each year in the United States. The emergence of broad-spectrum antibiotic resistance, particularly to methicillin, is a significant public health concern. S. aureus has become one of the most problematic infections in the hospital environment and, more recently, in the community setting. Consequently, large amounts of time and money have been invested to develop a vaccine against S. aureus, and while several candidates have been evaluated in human clinical trials, no program has successfully translated animal model efficacy data into an effective human vaccine. The goal of this proposal is to develop a broad-spectrum, multivalent subunit vaccine based on staphylococcal iron receptor proteins (SIRPs). Iron is an essential nutrient for bacteria, and SIRPs make excellent vaccine targets because they are highly conserved and are rapidly induced during infection in response to the low availability of iron in the host. A combination of four SIRPs, all natural membrane-bound lipoproteins in native S. aureus, were previously identified as immunogenic and efficacious in a model of mouse sepsis when administered as recombinants without acylation. The proposed studies will extend this work to a new, clinically relevant target indication of skin and soft tissue infections (SSTI). Unlike sepsis, protective responses against staphylococcal SSTI in mice appear to be concordant with those in humans in that they require a Th17-directed immune response. As lipoprotein-dependent activation of Toll-like receptor 2 (TLR2) is known to induce Th17 responses, we propose to immunize with acylated versions of the SIRP antigens, thereby returning them to their natural lipoprotein form and maximizing their potential to induce Th17 immunity through co-delivery of antigen and TLR2 agonist. The primary goal of this proposal is to measure the impact of acylation on vaccine efficacy in the murine SSTI model and identify a clinically appropriate adjuvant that optimizes efficacy and can be used in future studies to support commercialization of the vaccine. Evaluation of antigen-specific Th17 cellular and antibody responses will be used to monitor the induction of Th17 immunity and determine correlates of protection. Successful completion of the work outlined in this proposal will provide proof of concept for further development of a novel, recombinant S. aureus SSTI vaccine based on iron receptor lipoproteins. Clarifying the effect of natural protein acylation on immunogenicity and protection is an innovative approach that will contribute fundamental new knowledge to the field of protein vaccines. Furthermore, if successful, the Th17-targeted SIRP vaccine may be highly effective for additional indications beyond SSTI such as the clearance of nasal colonization and pulmonary infection in humans and mice, which have recently been shown to be Th17-dependent.